In various biological analyses, assay methods for detecting a target substance using a specific detectable label have been developed so far. For example, in the nucleic acid detection method by hybridization using the labelled nucleic acid, a nucleic acid (DNA or RNA) to be used as a probe is labelled and brought into contact with a sample containing a nucleic acid to be detected under conditions that hybrid can be formed. If the sample contains a nucleic acid having a base sequence complementary to that of the nucleic acid used as a probe, this nucleic acid binds (hybridize) to the probe to form a nucleic acid-nucleic acid hybrid. The target nucleic acid can be detected by measuring the label contained in the hybrid. In the immunoassay using labelled antigen or a labelled antibody, when an antigen is to be detected, it can be detected by labeling an antibody which is specifically bound to the antigen, effecting formation of an antigen-antibody complex, and detecting the label contained in the complex.
As the label to be used in such nucleic acid detection methods and immunoassay, radioactive substances, non-radioactive substances such as biotin or digoxigenin compounds, fluorescent substances, and the like are exemplified.
The following methods are known as the methods of introducing a fluorescent substance into, for example, a nucleic acid: the method of introducing a fluorescent substance-bound nucleotide into a nucleic acid using enzyme and the like (JP-A-6-271599); the method of binding a fluorescent substance-bound streptoavidin to a biotin-labelled nucleic acid; the method of reacting a fluorescent substance containing a group reactive with amine with an amino linker-bound nucleic acid; etc. However, these methods have such disadvantages that a fluorescent substance cannot be introduced into a naturally-occurring nucleic acid and complicated operation is required.
For example, JP-A-4-27867 describes a specific method of covalently binding a fluorescent rare earth metal chelate to a biological reactant, but this method is disadvantageous in that such a fluorescent rare earth metal chelate cannot be directly bound to a naturally-occurring nucleic acid via a covalent bond.
JP-A-6-94720 describes a method for photochemical labeling using a compound having a photoreactive group and a fluorescent rare earth metal chelate. However, this method is also disadvantageous in requiring a complicated step of ultraviolet irradiation.
Further, there is a method of intercalating a fluorescent substance to a double-stranded nucleic acid, which cannot be applied to labeling of a single-stranded nucleic acid.